فهرست مطالب bahman bahram nejad

Exendin-4 (Ex4) is a 39-amino acid peptide isolated from the salivary secretions of the lizard Heloderma susceptum and it has similar biological function as glucan-like peptide (GLP-1) receptor. The most important physiological role of GLP-1 is to regulate the amount of glucose metabolism by reducing the entry of glucose into the cells. Transgenic plants can be used as factories for the production of recombinant proteins. In order to maximize the efficiency of transgenic plants, the expression of foreign genes can be regulated in terms of time and place under the regulation of different promoters. Considering the nutritional value of carrot tuber and its fresh consumption, in this study, CTBEx4 gene was expressed in carrot plant using a root-specific promoter. In this study, in order to specifically express the CTBEx4 gene in carrot root, the Major Latex-Like promoter (MLL) related to the specific expression in sugar beet storage root tissue was replaced by the CaMV35S promoter in the pBI121 construct and were used for the transformation of carrot explants.

Using the CTAB method, DNA was extracted from the leaves of the sugar beet plant, then the PCR reaction was performed using specific MLL primers. The obtained fragment was replaced by the CaMV35S promoter in the pBI121 construct using restriction enzymes. Both pBI121-Mll-CTBEx4 and pBI121-CamV35S-CTBEx4 constructs were used to transform carrot leaf explants using Agrobacterium tumefaciens bacteria. Transgenic explants were indirectly regenerated into plants in MS culture media in vitro and then transferred to pot. Finally, the CTBEx4 gene expression of the transgenic plants was analyzed at mRNA and protein levels using RT-PCR and ELISA.

The results of the expression analysis using RT-PCR and ELISA methods indicated that the expression of CTBEx4 gene in carrot root tissue was regulated by MLL promoter, while no expression was observed in leaf tissue. Also, the results showed that the CTBEx4 gene was expressed under CaMV35S promoter regulation in both root and leaf tissues, but its expression in roots was lower (30%) than that under MLL promoter regulation. The obtained results indicated the ability of the MLL promoter to specific expression the CTBEx4 gene in the roots of carrot plants.

The MLL promoter was able to express the CTBEx4 gene in the root tissue of carrot plants at the mRNA and protein level, Also, CTBEx4 protein under MLL promoter had higher expression level compared to CaMV35S promoter.

Ferula pseudalliacea is a perennial and monocarpic plant species belonging to Apiace-ae family. This species is distributed from Middle Asia to Northern Europe. The main secondary me-tabolites present in Ferula genus comprise coumarins, sesquiterpene coumarins and di-sesquiterpene coumarins. Based on phytochemicals present in this plant species, it has been extensively utilized by local people in traditional medicine for long time. Phenylpropanoid pathway is mainly responsible for biosynthesis of these valuable phytochemicals. Cynamate-4-hydrolxylase (C4H) is one of the main and key enzyme in phenylpropanoid pathway. To date, there is no genomic or transcriptomic information in Ferula pseudalliacea and as the first report, we aimed to clone and sequence C4H and its expression analysis in root, stem, leaf, inflorescence and immature seed in this valuable medicinal plant species.

Different organs of F. pseudalliacea were collected from Gazne village near Sanandaj city in June 2019. RNA was extracted from collected organs using LiCl method. cDNA was synthesized using Yekta Tajihz Azmia kit in 20 µl reactions. PCR products amplified with pri-mer designed from conserved regions of C4H among apiaceous species and it was then cloned within pTG19. The verified recombinant plasmid were send for Applied Biosystem company for sequenc-ing. To confirm and asses the expression of C4H, both Real-Time PCR and semi-quantitative PCR were employed. The phylogenetic tree of C4H was obtained using Omega software based on the Neighbor-Joining method with bootstrap 1000 for amino acid sequence.

The obtained sequences from C4H clones was aligned and verified using BlastX. In the re-sulted dendrogram, C4H sequence from F. pseudalliacea was grouped with C4H gene from other species within Apiaceae family showing their clos relationship. The highest sequence identity was observed with C4H from Daucus carota which was fallen in same cluster with C4H from F. pseudal-liacea. It is shown that C4H has 7 motifs one of this motifs belongs to conserved motif cytochrome-cysteine 450 with FGVGRRSCPG sequence.

As the seconf most important gene in phenylpropanoid biosynthetic pathway, C4H plays an undeniable role in production of phenolic phytochemicals in particular in inflorescence which had the highest C4H expression rate. Studies performed on the C4H expression in other spe-cies revealed a specific action of C4H in biosynthesis of special secondary metabolites in specific tis-sue. As the phenolic compounds encompass a diverse range of secondary metabolites in forms of volatile and extracts, further investigations are needed to understand the exact role of C4H enzyme. Moreover, the identified gene sequence can be sued in production of recombinant constituents in other species which have the corresponding precursors.

Salinity is one of the environmental stresses that affects bread wheat grain yield in most parts of the world. One of the basic strategies to mitgiate the effect of non-biological stresses such as salinity is genetic improvement of crop plants. Identification of stress-associated genes is a prerequisite for genetic improvement. In the present study, the role of a number of genes in the aquaporin family and genes associated with the CDPK pathway in response to environmental stresses was identified as their expression profile analysis in response to salinity stress would facilitate breeding for salinity tolerance. In 2014, the expression of two genes TaNIP and TaSC was studied in two bread wheat cultivars "Bam" tolerant to salinity and "Tajan" sensitive to salinity under salinity stress (170 mM NaCl) and Non-saline conditions. The results showed that TaNIP expression in salt-tolerant cultivar (Bam) significantly increaseed in both leaf and root tissues under salinity stress conditions, but in sensitive cultivar (Tajan) the level of TaNIP expression, especially in leaf tissues, was lower and showed a decreasing trend. In cv. Bam in both leaf and root tissues, after 24 hours, TaSC expression increased in comparion to control. In cv. Tajan, the expression in leaf tissue was generally lower and after 24 hours, the expression level significantly decreased in comparion to control (non-saline). Moreover, expression analysis under the simultaneousapplication of salicylic acid andsodium chloride showed that the expression of both genes in leaf tissues of both bread wheat cultivars significantly increased and the gene expression showed an increasing trend. In conclusion, the results of this experiment indicated that the TaNIP and TaSC genes had different expression trends in tolerant and sensitive bread wheat cultivars under salinity stress conditions. Using salicylic acid as an elicitor also activated the hormonal transduction pathway, which induced the expression of both genes. These findings highlights the role of these genes in salinity tolerance of tolerant bread wheat cv. “Bam”.

Genetically improvement for salinity resistance is one of the most effective breeding methods for wheat. But understanding the mechanism, assessing and identifying salinity tolerance genes is a prerequisite for genetic modification. The role of a number of families of transcription factors has been proven in response to environmental stresses, hence, their identification and investigation facilitates the possibility of overcoming stresses. Considering the significant role of the important family of WRKY transcription factors in response to stresses, the expression of two important genes of this family including TaWRKY10 and TaWRKY53 in different tolerant and susceptible cultivars of wheat was investigated. The results showed that TaWRKY10 exhibited different expressions in susceptible wheat (Tajan) and resistant (Bam) cultivars. In resistant cultivar (Bam) under saline conditions over a period of 48 hours, TaWRKY10 showed more expression level in both leaf and root tissues than the control and also the sensitive (Tajan) cultivar. However, the expression of TaWRKY53 was different in resistant and susceptible cultivars. TaWRKY53 in susceptible cultivar (Tajan), over the period of 48 hours, showed more expression level than the control and resistant cultivar (Bam) in leaves and root tissues. Also, by using salicylic acid under salinity stress, the expressions of TaWRKY10 and TaWRKY53 in Tajan and Bam cultivars were different. In general, our results show that WRKY transcription factors can play a role in generating resistance to salinity stress in wheat and therefore can be used in genetic improvement.

Endophytes are microorganisms (bacteria or fungi or actinomycetes) that reside inside the internal tissues of plant with no negative effect on host plants. Endophytic bacteria trigger many positive effects on host plants. The aroma and flavor of strawberry fruit are important characteristics that is affected by endophytic bacteria such as Methylobacterium. Endophytic bacteria, are effective in the synthesis of carotenoids and the production of volatile and phenolic compounds. In addition, they are effective in producing the aromatic components in fruit. Endophytic bacterial frequency and genotype can be affected by host plant genotype, climatic and soil conditions. This study was carried out to isolate endophytic bacteria from two strawberry cultivars (Kurdistan and Parous) in three regions (Angouzan, Gavrood and Noshur) in Kurdistan province. For accurate identification of bacterial isolates, 16S-rRNA gene sequencing was used for identification of purified endophytic isolates and their frequency determined at different times of fruit ripening. The results revealed that the all endophytic bacterial isolates from both cultivars were belong to the genus Methylobacterium. The frequency of these bacteria was statistically different in two cultivars and in different sampling time. The frequency of Methylobacterium significantly was higher in Kurdistan cultivar compared to Parous cultivar in all three regions (p <0.05). The highest frequency of Methylobacterium was observed in the fruit ripening time. It seems that Methylobacterium and its frequency probably have an effect on the aroma and delicacy of these two varieties.

Wheat (Triticum aestivum L.) is one of the most important grain products in the world which is the main food of the people in many parts of the world and provides about 20% of the calories needed by the world population. The late stages of wheat growth in the Mediterranean regions are usually accompanied by drought stress. Supplemental irrigation is one of the appropriate strategies to reduce the damaging effects of drought stress and improve yield and sustainability. Nitrogen is the most important nutrient in rainfed farming after wheat planting is the most important tool available to increase yield. Nitrogen fertilizer application to soil at late vegetative stage may not be appropriate due to dry soil surface and reduced root activity, Therefore Nitrogen spraying can be used as a quick and efficient way to meet plant nutrition due to its advantages such as faster and more uptake by the plant.

In order to study the effect of supplemental irrigation and different rates of urea fertilizer and urea foliar application on dry matter remobilization, yield and yield components and Grain protein contents experimental wheat in split –split plot in randomized complete block design with three replications It was designed and implemented in the research farm of Kurdistan University in 2016-2017. Treatments included irrigation levels (no irrigation and irrigation at booting stage) as main factor, three dry wheat cultivars (Rejaw, Sardari and Azar 2) as sub factor of different rates of nitrogen fertilizer (50 kg urea in fall (A), A + 50 kg urea in the spring (B) and B + foliar solution (20 kg urea in the booting stage) were used as a sub-factor. The studied traits included leaf dry matter, stem + leaf pod and spike in flowering and ripening stages, leaf dry matter remobilization, stem + leaf pod and panicle seed, total soil remobilized dry matter, soil yield. Dry leaf remobilization, stem + leaf pod and pod straw, total remobilization efficiency, remobilization share in grain yield, spike number per square meter, spike number per grain, 1000-grain weight, biological yield, yield, harvest index and grain protein content.Leaf dry matter, stem + Sheath dry matter and spike dry matter in flowering and maturity stages, leaf dry matter remobilization, stem + Sheath dry matter, Chaff dry matter, total dry matter remobilized, leaf,, stem + sheath, chaff and total dry matter remobilization efficiency, remobilization share in grain yield, spike number per square meter, kernals per spike, 1000-grain weight, biological yield, yield, harvest index and grain protein content were measured.                                                        

Results showed that the amount of stem + sheath, dry matter at flowering and maturity, leaf, stem + sheath, spike remobilization dry matter, total remobilization, stem+ sheath remobilization efficiency, remobilization share in grain yield, weight Spike, kernals per spike, 1000-grain weight, yield and biological yield were affected by supplemental irrigation.Among cultivars in terms of stem + sheath and chaff dry matter at flowering and maturity, leaf, stem + Sheath, spike remobilized dry matter, total remobilization, leaf remobilization efficiency, remobilization share in grain yield, weight Spike, kernals per spike, 1000-grain weight, yield, biological yield, harvest index and grain protein were different. Rejaw cultivar had more grain number per spike and yield than Azar2 and Sardari.Different rates of nitrogen fertilizer had a significant effect on stem + sheath, spike and total dry matter remobilization efficiency and remobilization share on grain yield.Supplemental irrigation at the booting stage increased dry matter stored in stem + sheath and increased the amount of remobilized dry matter during grain filling.Irrigation at booting stage increased grain yield by 12.4%. Increasing urea fertilizer and urea foliar application at the end of growth stages increased plant yield and grain quality by providing more nitrogen and stay-green.

In this study, three nonpathogenic endophytic bacterial isolates recovered from Fragaria × ananassa stolon and petal and their antagonistic activity was evaluated against Colletotrichum nymphaeae, the causal agent of strawberry anthracnose under in vitro, in vivo, and greenhouse conditions. Bacterial isolates were identified as Bacillus spp. using a combination of phenotypic, biochemical properties and molecular phylogenetic analysis of the 16S rDNA gene sequence. The living cells of bacterial isolates inhibited the mycelial growth of C. nymphaeae using dual culture method and inhibition percentage was evaluated as 41.87, 50, and 54.92 % in MarG2, MarD40 and MarD35, respectively. The volatile compounds (VOCs) produced by bacterial isolates inhibited the mycelial growth of C. nymphaeae and the highest inhibition was observed in MarD35 isolate (24.7%). Moreover, the cell free culture filtrates of bacterial isolates reduced the mycelial growth and conidial germination of pathogen and the highest inhibition was observed in MarG2 (34.96%) on mycelial growth and in MarD35 (52.07%) on conidial germination. The living cells of bacterial isolates decreased anthracnose fruit rot at postharvest and the highest inhibition was observed in MarD40 (85.63%) and MarD35 (81.72%), respectively. Furthermore, disease severity of strawberry anthracnose by bacterial isolates was reduced using drenching soil and plant spraying techniques. The findings of this study showed that bacterial isolates have the ability to prevent the growth of fungal pathogen under in vitro, in vivo and greenhouse conditions.

Resveratrol is an antioxidant secondary metabolite which belongs to a specific phytoalexins called stilbenes. Grape has been considered as the main source of resveratrol in the human diet. Many studies have been conducted on genus Vitis due to the presence of high levels of polyphenolic compounds in different tissues. In recent years, a lot of effort has been made to increase resveratrol yield in Vitis vinifera via tissue and hairy roots culture. In the present study, hairy roots have been produced by Agrobacterium rhizogenes strain ATCC15834 and successfully produced resveratrol in internode of grape genotype W16. Transformations of hairy roots were recognized by PCR using specific primers of rolB gene. After stimulation, the effect of abiotic elicitors with different concentrations including methyl jasmonate, sodium acetate, acetic acid and ammonium nitrate were tested on the production of hairy root biomass and resveratrol. Resveratrol content was measured by TLC and HPLC methods. Results showed that the capacity of hairy roots for resveratrol production is higher than natural roots. A significant difference was observed between different elicitors in terms of hairy root biomass and resveratrol amount. Findings revealed that treatment with 3 mM acetic acid and 50 μM methyl jasmonate led to the highest and lowest amount of hairy roots biomass and resveratrol content, respectively. According to the fidings of the present study it can be stated that both hairy roots and different elicitors are effective in biomass and resveratrol production. This method can be used to increase the yield of resveratrol for large scale production via tissue culture.

The purpose of this research was to transform two economically important cultivars of strawberry with P5CS gene، which encodes Δ1-pyrroline-5-carboxylate synthetase (P5CS)، the key enzyme in proline biosynthesis. Shoots were obtained on MS basal medium supplemented with 2% glucose and 4 mg/l TDZ for Camarosa and Kurdistan cultivars. For genetic transformation، a binary vector PBI121 containing P5CS gene under control of the 35SCaMV promoter was used. Transformed cells (explants) were regenerated on the selective regeneration medium containing 75 mg/l kanamycin and 500 mg/l cefotaxime after five days of pre-incubation and 72 h of co-cultivation with Agrobacterium، while control explants failed to grow in the same medium. The presence of the transgene in the plant genome was confirmed by PCR. The morphology of the transgenic plants was normal as controls. Drought stress was applied using polyethylene glycol (PEG) 6000 at concentrations equal to 0، -5 and -7 Bar، respectively. Proline content was four times higher in the transformed leaves compared to that of the untransformed plants while the proline content in the roots was similar in both transgenic and wild-type plants. Overproduction of P5CS also increased chlorophyll content، shoot length، shoot fresh and dry weight in the transgenic plants under drought-stress conditions.

One of the most important proteins in gene silencing and creation of small RNA، are argonaute proteins. Most of these proteins have an endonucleolytic activity so argonaute proteins play a key role in creation of defense mechanism against certain pathogenic viruses. In current study we studied the putative argonaute genes in six medicinal plants using bioinformatic tools. Sequences of argonaute genes of six medicinal plants were retrieved from target database using Arabidopsis Argonaut blast. Sequence alignment، 3D structure، phylogenetic tree and conserved domain were generated for six medicinal plants argonautes. Results showed that studied medicinal plants have 6 to 18 argonaute proteins in the genome. There were three classes of Argonaute proteins similar to that of Arabidopsis، and each varies in length from 846 to 1187 AA، with a average of 970 AA and 108 kDa. six medicinal plants argonaute proteins had PAZ، PIWI and MID conserved domains. Our results showed that six medicinal plants have three classes of argonautes، This indicates is that the conservation of argonaute proteins in different plants.

In order to optimize callus induction in Pistacia atlantica Desf. subsp. kurdica Rech. f., the effect of different plant growth regulator (PGR) combinations were studied. Inflorescence axis explants were cultured on WPM medium supplemented with 1, 2 and 3 mg/L of TDZ or 1, 2 and 3 mg/L of BA, in combination with 1mg/l different auxins: 2, 4-D, NAA and IBA . Callus initiation was observed in all media evaluated and the highest percentage (100%) was observed in medium containing 1 mg/L BA or TDZ and 1 mg/L NAA. Additionally, anthocyanin, total phenolic compound of callus and immature fruits were measured. Anthocyanin content of callus and immature fruits were 79.42 ± 4.63 and 61.71 ± 3.81 mg cyanidin-3 glucoside/g fresh weights, respectively. Total phenolic content of callus and fruits were 4.91 ± 0.492 and 5.1 ± 0.780 mg gallic acid/g fresh weights. GC-MS analysis of essential oil constituents of P. atlantica subsp. kurdica callus and leaves showed 8 kinds of substances in callus and 13 in leaves. Main composition of callus extract were alpha pinene (89.19%), camphene (1.09%) and Bicyclo[3.1.1] heptane, 6,6-dimethyl-2-methylene (3.40%) and it was alpha Pinene (15.63%), gamma elemene (33.84%) and caryophyllene (9.26%) in leaves.

P. atlantica subsp. Kurdica, with the local name of Baneh, is a wild medicinal plantwhich grows in Kurdistan, Iran. The identification of resistance gene analogs holdsgreat promise for the development of resistant cultivars. A PCR approach withdegenerate primers designed according to conserved NBS-LRR (nucleotide bindingsite-leucine rich repeat) regions of known disease-resistance (R) genes was used toamplify and clone homologous sequences from P. atlantica subsp. Kurdica. A DNAfragment of the expected 500-bp size was amplified. The nucleotide sequence of thisamplicon was obtained through sequencing and the predicted amino acid sequencecompared to the amino acid sequences of known R-genes revealed significant sequencesimilarity. Alignment of the deduced amino acid sequence of P. atlantica subsp.Kurdica resistance gene analog (RGA) showed strong identity, ranging from 68% to77%, to the non-toll interleukin receptor (non-TIR) R-gene subfamily from other plants.A P-loop motif (GMMGGEGKTT), a conserved and hydrophobic motif GLPLAL, akinase-2a motif (LLVLDDV), when replaced by IAVFDDI in PAKRGA1 and a kinase-3a (FGPGSRIII) were presented in all RGA. A phylogenetic tree, based on the deducedamino-acid sequences of PAKRGA1 and RGAs from different species indicated thatthey were separated in two clusters, PAKRGA1 being on cluster II. The isolated NBSanalogs can be eventually used as guidelines to isolate numerous R-genes in Pistachio.

Genetic transformation studies were carried out to standardize a protocol for Agrobacterium -mediated genetic transformation of three economically important strawberry (Fragaria x ananassa Duch) cultivars Kurdistan’، Camarosa’، and ‘Paros’. Shoot regeneration frequency 72، 65 and 30% was obtained on MS (1) basal medium supplemented with 2% glucose and 4 mg/l TDZ for Camarosa، Kurdistan and Paros، respectively. To optimize the concentration of kanamycin for these cultivars، we observed the responses of leaf explants to different concentration of kanamycin (0-100 mg/l) in the media. Our results showed that 75 mg/l was an effective concentration of kanamycin. For genetic transformation، Agrobacterium tumefaciens strain C58C1RifR (pGV2260) harbouring the binary vector pTJK136 containing uidA gene with intron along with kanamycin resistance gene (npt-II) was used. After five days pre-incubation and 72 h co-cultivation، transformed cells (explants) were able to grow on the selective regeneration medium containing 75 mg/l kanamycin and 500 mg/l cefotaxime، while، control explants failed to grow. The regenerated putative transgenic shoots were analyzed by histochemical GUS assay and PCR analysis. Transformation efficiency based on inoculated explants number was 2، 1 and 3% for ‘، Camarosa’، ‘Paros ’، and ‘Kurdistan’، respectively. The standardized protocol would be useful for Agrobacterium-mediated genetic transformation of these cultivars with important agronomic genes.

In order to investigate the responses of Sardari wheat ecotypes to drought stress, 12 spikes of Sardari wheat were selected from different regions of Kurdistan province and planted as head to row. In the second year, a split- plot factorial field experiment was conducted. In this experiment water stress and irrigation were considered as the levels of main factor and 12 Sardari wheat ecotypes allocated to subplots. In the third year, the harvested wheat ecotypes in the second year were grown under control and drought conditions to evaluate grain yield and stress susceptibility index (SSI). Results showed that photosynthesis rate, rate of leaf water loss, protein and chlorophyll content, mesophyll conductance, photosynthetically water use efficiency and sub stomatal CO2 concentration were affected significantly by water stress in all wheat landraces. The results revealed that ecotypes with higher photosynthetic rate showed higher transpiration. These ecotypes exhibited better mesophyll conductance and lower SSI. Leaf protein patterns of ecotypes by electrophoresis were similar to that of soluble protein content under drought stress. The results showed that under drought stress, the new bands of protein were identified which correlated with pattern of photosynthesis of ecotypes in this condition.

Genetic transformation studies were carried out to standardize a protocol for Agrobacterium -mediated genetic transformation of three economically important strawberry (Fragaria x ananassa Duch) cultivars Kurdistan’, Camarosa’, and ‘Paros’. Shoot regeneration frequency 72, 65 and 30% was obtained on MS (1) basal medium supplemented with 2% glucose and 4 mg/l TDZ for Camarosa, Kurdistan and Paros, respectively. To optimize the concentration of kanamycin for these cultivars, we observed the responses of leaf explants to different concentration of kanamycin (0-100 mg/l) in the media. Our results showed that 75 mg/l was an effective concentration of kanamycin. For genetic transformation, Agrobacterium tumefaciens strain C58C1RifR (pGV2260) harbouring the binary vector pTJK136 containing uidA gene with intron along with kanamycin resistance gene (npt-II) was used. After five days pre-incubation and 72 h co-cultivation, transformed cells (explants) were able to grow on the selective regeneration medium containing 75 mg/l kanamycin and 500 mg/l cefotaxime, while, control explants failed to grow. The regenerated putative transgenic shoots were analyzed by histochemical GUS assay and PCR analysis. Transformation efficiency based on inoculated explants number was 2, 1 and 3% for ‘, Camarosa’, ‘Paros ’, and ‘Kurdistan’, respectively. The standardized protocol would be useful for Agrobacterium-mediated genetic transformation of these cultivars with important agronomic genes.